scholarly journals A Gene Expression Signature to Predict Nucleotide Excision Repair Defects and Novel Therapeutic Approaches

2021 ◽  
Vol 22 (9) ◽  
pp. 5008
Author(s):  
Rongbin Wei ◽  
Hui Dai ◽  
Jing Zhang ◽  
David J. H. Shih ◽  
Yulong Liang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Nucleotide Excision Repair ◽  
Cell Lines ◽  
Excision Repair ◽  
Gene Expression Signature ◽  
Gene Signature ◽  
Expression Signature ◽  
Nucleotide Excision ◽  
Functionally Impaired ◽  
Synthetically Lethal

Nucleotide excision repair (NER) resolves DNA adducts, such as those caused by ultraviolet light. Deficient NER (dNER) results in a higher mutation rate that can predispose to cancer development and premature ageing phenotypes. Here, we used isogenic dNER model cell lines to establish a gene expression signature that can accurately predict functional NER capacity in both cell lines and patient samples. Critically, none of the identified NER deficient cell lines harbored mutations in any NER genes, suggesting that the prevalence of NER defects may currently be underestimated. Identification of compounds that induce the dNER gene expression signature led to the discovery that NER can be functionally impaired by GSK3 inhibition, leading to synergy when combined with cisplatin treatment. Furthermore, we predicted and validated multiple novel drugs that are synthetically lethal with NER defects using the dNER gene signature as a drug discovery platform. Taken together, our work provides a dynamic predictor of NER function that may be applied for therapeutic stratification as well as development of novel biological insights in human tumors.

Identification of a Novel Gene Expression Signature in Mantle Cell Lymphoma from the Fondazione Italiana Linfomi (FIL)-MCL-0208 Trial: A Focus on the B Cell Receptor Pathway

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 701-701
Author(s):  
Riccardo Bomben ◽  
Simone Ferrero ◽  
Michele Dal Bo ◽  
Tiziana D'Agaro ◽  
Alessandro Re ◽  
...  
Keyword(s):  
Gene Expression ◽  
Board Of Directors ◽  
Cell Lines ◽  
Gene Expression Signature ◽  
Gene Signature ◽  
Advisory Committees ◽  
Expression Signature ◽  
Qrt Pcr ◽  
Mantle Cell ◽  
Bcr Signaling

Abstract Background. The aggressive clinical behavior of mantle cell lymphoma (MCL) is attributed to specific genetic and molecular mechanisms involved in its pathogenesis, mainly the t(11;14)(q13;q32) traslocation and cyclin D1 (CCND1) overexpression. Nevertheless, evidence of a certain degree of clinical/biological heterogeneity has been disclosed by gene expression profile (GEP) and (immuno)genetic/immunohistochemistry studies. Aim. To use a GEP approach to identify MCL subsets with peculiar clinical/biological features in the context of MCL patients treated homogeneously with an autologous transplantation-based program. Methods. The study was based on a cohort of 42 MCL cases enrolled in the Fondazione Italiana Linfomi (FIL)-MCL-0208 randomized Italian clinical trial. Purified clonal CD19+ MCL cells were obtained by high-speed cell sorting of peripheral blood MCL samples. GEP experiments were performed in 30 cases, with Agilent platform. Bioinformatics analyses were performed by Gene Springs and Gene Set Enrichment Analysis (GSEA) software. Gene signature validations were performed by quantitative real time PCR (QRT-PCR). Results. i)Unsupervised and supervised analyses. Unsupervised analysis by principal component analysis (PCA) was able to divide the cohort in two main subgroups named PCA1 (12 cases) and PCA2 (18 cases). Supervised analysis by segregating cases according to the PCA1 and PCA2 classification defined a gene expression signature of 710 gene (234 up-regulated) that highlighted a constitutive overexpression of genes of the BCR signaling pathway. Consistently,GSEA showed a significant enrichment of genes belonging to 3 gene sets related to BCR signaling. ii) Identification of a "PCA2-type" gene signature. By merging the list of differentially expressed genes according to supervised analysis of GEP data and the gene list related to BCR signaling according to GSEA, a group of 9 genes, all overexpressed in PCA2 cases, i.e. AKT3, BLNK, BTK, CD79B, PIK3CD, SYK, BCL2, CD72, FCGR2B, was obtained. Among these genes, a subgroup of 6 genes, i.e. AKT3, BLNK, BTK, CD79B, PIK3CD, SYK, was selected for the direct involvement in the BCR pathway, and utilized for further validations. iii) Generation of a 6-gene prediction model. The selected 6 genes were then utilized to generate a prediction model by using 20 cases as training sub-cohort and the remaining 10 cases as validation cohort. By this approach, 9/10 cases of the validation cohort were correctly assigned according to the PCA2/PCA1 classification. The model was re-tested by QRT-PCR in 24 cases used in the GEP (16 for training and 8 for validation), and again, 7/8 cases of the validation sub-cohort were correctly classified. QRT-PCR was then utilized to classify further 12 cases (7 cases defined as PCA2) not employed for GEP analysis. Overall, in the 42 cases, 23 cases were considered as PCA2 with the GEP/QRT-PCR approach. iv) Clinical/biological correlations. No association was found between the 6-gene signature and IGHV status (22/30 unmutated IGHV cases) or between the signature and the overexpression of SOX11 (17/30 cases over the median value). In addition, no association was found with the presence of the main recurrent mutations of the ATM, BIRC3, CCND1, KMTD2, NOTCH1, TP53, TRAF2, WHSC1 genes. Finally, an "ad-interim" analysis of progression free survivals (PFS) (Cortelazzo et al EHA, 2015) suggested a trend for a shorter PFS (2-years PFS 45% vs 72%, p=0.08) for cases classified as PCA2 by the GEP/QRT-PCR approach. v) 6-gene signature and sensitivity to the BCR inhibitor ibrutinib. The finding that PCA2 cases overexpressed BCR-related genes and had a more aggressive clinical course prompted us to investigate the 6-gene signature in the context of ibrutinib sensitive/resistant MCL cell lines. To do this, the proliferation rate of the MCL cell lines REC1, JEKO1, UPN1, GRANTA, JVM2, Z138 was investigated either in presence or in absence of ibrutinib 10 nanoM for 7 days. REC1, JEKO1 were selected as responsive by showing ≥80% inhibition upon ibrutinib. Of note, responsive cell lines showed higher expression levels of the 6-gene signature then the resistant counterpart, as evaluated by QRT-PCR. Conclusions. A novel 6-gene expression signature related to the BCR pathway has been found to characterize MCL cells with peculiar clinical/biological features and sensitivity to BCR inhibitors. Disclosures Luminari: Roche: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Teva: Membership on an entity's Board of Directors or advisory committees.

scholarly journals Nucleotide Excision Repair (NER) Is Frequently Impaired and Affects Outcome in Multiple Myeloma (MM)

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 2055-2055
Author(s):  
Raphael Szalat ◽  
Matija Dreze ◽  
Mehmet Kemal Samur ◽  
Anne S. Calkins ◽  
Giovanni Parmigiani ◽  
...  
Keyword(s):  
Gene Expression ◽  
Multiple Myeloma ◽  
Dna Repair ◽  
Genomic Instability ◽  
Nucleotide Excision Repair ◽  
Cell Lines ◽  
Excision Repair ◽  
Sequencing Data ◽  
Nucleotide Excision

Abstract Introduction Multiple Myeloma (MM) is a heterogeneous disease characterized by genomic instability and eventual poor outcome. Aberrations in DNA repair-related pathways have been considered to explain the instability. Nucleotide excision repair (NER) is an important pathway involved in the removal of bulky adducts and DNA crosslinks induced by various genotoxins. Little is known about the relationship between NER in MM biology and patient outcomes. Here we assess the role of NER in MM. Methods We evaluated NER efficiency in a panel of MM cell lines (n=18), with a functional assay based on the purified DNA-Damage Binding protein 2 (DDB2) complex (DDB2 proteo-probe, Dreze et al. 2014). NER proficiency was correlated with cytogenetic characteristics, p53 status, sequencing data, gene expression profile, and with melphalan (MLP) sensitivity evaluated by CellTiterGlo (CTG). We then evaluated NER efficiency in patient samples and interrogated the role of NER in MM patients by correlating expression of NER genes with survival (OS) in a cohort of 170 patients (IFM 2005-01) homogeneously treated with alkylating agents. Results NER, measured as the amount of (6-4) photoproducts remaining 2 hours after UV irradiation, showed variability between MM cell lines. Out of 18 cell lines, 7 exhibited various levels of NER deficiencies, defined as less than 90% repair at 2 hours (4 cell lines 90-70% and 3 cell lines <60%). The other 11 cell lines presented more than 90% of repair. P53 loss of function did not associate with NER deficiency. Notably, all t(4;14) cell lines tested (n=5) showed a NER repair rate > 90%. NER deficient cell lines (NER <90%) were sensitive to melphalan. However all melphalan sensitive cells did not exhibit NER deficiency, This suggests that other DNA repair pathways are involved in the repair of melphalan-induced lesions. Furthermore, we performed the assay in patient samples showing variable levels of NER, which may reflect different disease status and prognosis. Whole genome sequencing data from 6 NER deficient cell lines revealed missense mutations in critical NER genes in 2 of these cell lines. MM1S and MM1R cells showed mutations in the Xeroderma Pigmentosum Complementation Group A (XPA) gene (mutation D70H), and MM1R was also mutated in the Cockayne syndrome, ERCC6 gene (mutation L682I). Gene expression profile comparison in 12 of these showed a positive correlation between expression of NER genes and NER efficiency. We next studied expression of 20 NER genes in 170 patients treated with high dose melphalan (IFM 2005-01). The analysis revealed a significant negative correlation between 5 overexpressed NER genes (ERCC3, ERCC4, ERCC6, MMS19 and NTHL1) and overall survival (OS). Conclusion NER efficiency is heterogeneous in MM, in part due to acquired mutations. Impairment of NER is associated with outcome as well as may contribute to genomic instability. Ability to proficiently measure NER in patient samples provides us an opportunity to now evaluate NER as a prognostic marker in myeloma. Disclosures No relevant conflicts of interest to declare.

scholarly journals Comparative analyses of two primate species diverged by more than 60 million years show different rates but similar distribution of genome-wide UV repair events

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Umit Akkose ◽  
Veysel Ogulcan Kaya ◽  
Laura Lindsey-Boltz ◽  
Zeynep Karagoz ◽  
Adam D. Brown ◽  
...  
Keyword(s):  
Nucleotide Excision Repair ◽  
Cell Lines ◽  
Human Cell ◽  
Excision Repair ◽  
Mouse Lemur ◽  
Repair Mechanism ◽  
Damage Repair ◽  
Genome Wide ◽  
Repair Efficiency ◽  
Nucleotide Excision

Abstract Background Nucleotide excision repair is the primary DNA repair mechanism that removes bulky DNA adducts such as UV-induced pyrimidine dimers. Correspondingly, genome-wide mapping of nucleotide excision repair with eXcision Repair sequencing (XR-seq), provides comprehensive profiling of DNA damage repair. A number of XR-seq experiments at a variety of conditions for different damage types revealed heterogenous repair in the human genome. Although human repair profiles were extensively studied, how repair maps vary between primates is yet to be investigated. Here, we characterized the genome-wide UV-induced damage repair in gray mouse lemur, Microcebus murinus, in comparison to human. Results We derived fibroblast cell lines from mouse lemur, exposed them to UV irradiation, and analyzed the repair events genome-wide using the XR-seq protocol. Mouse lemur repair profiles were analyzed in comparison to the equivalent human fibroblast datasets. We found that overall UV sensitivity, repair efficiency, and transcription-coupled repair levels differ between the two primates. Despite this, comparative analysis of human and mouse lemur fibroblasts revealed that genome-wide repair profiles of the homologous regions are highly correlated, and this correlation is stronger for highly expressed genes. With the inclusion of an additional XR-seq sample derived from another human cell line in the analysis, we found that fibroblasts of the two primates repair UV-induced DNA lesions in a more similar pattern than two distinct human cell lines do. Conclusion Our results suggest that mouse lemurs and humans, and possibly primates in general, share a homologous repair mechanism as well as genomic variance distribution, albeit with their variable repair efficiency. This result also emphasizes the deep homologies of individual tissue types across the eukaryotic phylogeny.

31 Effect of vitrification on global gene expression dynamics of bovine elongating embryos

2020 ◽  
Vol 32 (2) ◽  
pp. 141
Author(s):  
Z. Jiang ◽  
E. Gutierrez ◽  
H. Ming ◽  
B. Foster ◽  
L. Gatenby ◽  
...  
Keyword(s):  
Gene Expression ◽  
Oxidative Phosphorylation ◽  
Mitochondrial Dysfunction ◽  
Nucleotide Excision Repair ◽  
Excision Repair ◽  
Cell Junction ◽  
Rna Seq ◽  
Protein Ubiquitination ◽  
Stage Of Development ◽  
Nucleotide Excision

The ability to cryopreserve gametes and embryos has been a valuable tool for reproductive management in all mammalian species, especially livestock. Embryo vitrification involves exposure to high concentrations of cryoprotectants and osmotic stress during cooling and warming. These factors have to affect gene expression. The elongating embryo is a stage of embryo development that can be recovered noninvasively in the cow on day (D) 14 and represents a critical stage of development when many embryos die. In this study, we aimed to evaluate the effect of vitrification on the transcriptome dynamics of D14 embryos by RNA sequencing (RNA-seq). Invitro blastocyst-stage embryos were vitrified by exposure to dimethyl sulfoxide and ethylene glycol solution, followed by placing on Cryo Loks and plunging in liquid nitrogen. After warming, embryos were loaded into straws and transferred into eight synchronized recipients, four cows received nonvitrified embryos and four cows received vitrified embryos (20 embryos per cow). Embryo flushing yielded 12 nonvitrified and 9 vitrified viable D14 embryos. Whole embryos (six nonvitrified and two vitrified embryos) or isolated trophectoderm (TE; four nonvitrified and seven vitrified) were processed for RNA-seq. The Smart-sEqn 2 protocol was followed to prepare RNA-seq libraries. Sequencing reads were prefiltered and aligned to the bovine genome, and gene expression values were calculated as fragments per kilobase of transcript per million mapped reads. Genes were deemed differentially expressed between treatments if they showed a false discovery rate P-value&lt;0.05 and fold-change &gt;2. Ingenuity pathway analysis was used to reveal gene ontology and pathways. Expression of 927 genes was changed in D14 embryos as a result of vitrification, with 782 and 145 genes upregulated and downregulated, respectively. In TE, vitrification resulted in 4096 and 280 upregulated or downregulated genes, respectively. Several pathways were upregulated by vitrification in both whole embryos and TE, including epithelial adherens junctions, sirtuin signalling, germ cell-Sertoli cell junction, ATM signalling, nucleotide excision repair, and protein ubiquitination pathways. Downregulated pathways included EIF2 signalling, oxidative phosphorylation, mitochondrial dysfunction, regulation of eIF4 and p70S6K signalling, mammalian target of rapamycin signalling, sirtuin singling, and nucleotide excision repair pathways. In addition, we found 671 and 61 genes upregulated and downregulated in both vitrified whole embryos and TE. Mitochondrial dysfunction and oxidative phosphorylation signalling were upregulated, whereas epithelial adherens junction and sirtuin signalling were downregulated, suggesting mitochondrial function and energy production were impaired in TE after vitrification. Our analysis identified specific pathways and implicated specific genes affected by cryopreservation and potentially affecting embryo developmental competence.

The Expression Signature of M3 AML Suggests Direct and Indirect Effects of PML-RARA on Target Genes.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 719-719 ◽  
Author(s):  
Jacqueline E. Payton ◽  
Nicole R. Grieselhuber ◽  
Li-Wei Chang ◽  
Mark A. Murakami ◽  
Wenlin Yuan ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
Cell Lines ◽  
Binding Sites ◽  
Expression Profiles ◽  
Gene Expression Signature ◽  
Myeloid Cell ◽  
Expression Signature ◽  
Signature Genes ◽  
Cell Fractions

Abstract In order to better understand the pathogenesis of acute promyelocytic leukemia (APL, FAB M3), we sought to determine its gene expression signature by comparing the expression profiles of 14 APL samples to that of other AML subtypes (M0, M1, M2, M4, n=62) and to fractionated normal whole bone marrow cells (CD34 cells, promyelocytes, PMNs, n=5 each). We used ANOVA and SAM (Significance Analysis of Microarrays) to select genes that were highly expressed in APL cells and that displayed low to no expression in other AML subtypes. The APL signature was then further refined by filtering genes whose expression in APL was not significantly different from that of normal promyelocytes, yielding 1121 annotated genes that reliably distinguish APL from the other FAB subtypes using unsupervised hierarchical clustering, both in training and validation datasets. Fold change differences in expression between M3 and other AML FAB classes were striking, for example: GABRE 35.4, HGF 21.3, ANXA8 21.3, PTPRG 16.9, PTGDS 12.1, PPARG 11.1, STAB1 9.8. A large proportion of the APL versus other FAB dysregulome was recapitulated when we compared APL expression to that of the normal pattern of myeloid development. We identified 733 annotated genes with significantly different expression in APL versus normal myeloid cell fractions. These dysregulated genes were assigned to 4 classes: persistently expressed CD34 cell-specific genes, repressed promyelocyte-specific genes, prematurely expressed neutrophil-specific genes and genes with high expression in APL and low/no expression in normal myeloid cell fractions. Expression differences in several of the most dysregulated genes were validated by qRT-PCR. We then examined the expression of the APL signature genes in myeloid cell lines and tumors from a murine APL model. The bona fide M3 signature was not apparent in resting NB4 cells (which contain t(15;17), and which express PML-RARA), nor in PR-9 cells following Zn induction of PML-RARA expression, suggesting that neither cell line accurately models the gene expression signature of primary APL cells. Most of the nodal genes of the mCG-PML-RARA murine APL dysregulome (Yuan, et al, 2007) are similarly dysregulated in human M3 cells; however, the human and mouse dysregulomes do not completely coincide. Finally, we have begun investigating which APL signature genes are direct transcriptional targets of PML-RARA. The promoters of the APL signature genes were analyzed for the presence of known PML-RARA binding sites using multiple computational methods. The analyses demonstrated that several transcription factors (EBF3, TWIST1, SIX3, PPARG) have putative retinoic acid response elements (RAREs) in their upstream regulatory regions. Additionally, we examined the promoters of some of the most upregulated genes (HGF, PTGDS, STAB1) for known consensus sites of these transcription factors, and found that all have putative binding sites for at least one. These results suggest that PML-RARA may initiate a transcriptional cascade that relies not only on its own activity, but also on the actions of downstream transcription factors. In summary, our studies indicate that primary APL cells have a gene expression signature that is consistent and highly reproducible, but different from commonly used human APL cell lines and a mouse model of APL. The molecular mechanisms that govern this unique signature are currently under investigation.

A gene expression signature to predict high-grade prostate cancer response to oxaliplatin.

2012 ◽  
Vol 30 (5_suppl) ◽  
pp. 150-150
Author(s):  
Philippe Pourquier ◽  
Stephane Puyo ◽  
Pierre Richaud ◽  
Jacques Robert ◽  
Nadine Houede
Keyword(s):  
Gene Expression ◽  
Prostate Cancer ◽  
Clinical Trials ◽  
Cell Lines ◽  
Gene Expression Signature ◽  
Chemotherapeutic Agents ◽  
Low Grade ◽  
Gleason Grade ◽  
High Grade ◽  
Expression Signature

150 Background: Prostate cancer (PCa) is one of the leading causes of death from cancer in men. High Gleason grade prostate cancers are characterized by aggressive tumors with poorly differentiated cells and a high metastatic potential. They are often refractory to chemical castration but still treated with hormone therapy to which docetaxel or cabazitaxel are added when they become resistant to the anti-androgen. Despite many clinical trials with other chemotherapeutic agents, response rates remain low. Moreover, none of these trials took into account the tumor grade. Methods: We used an in silico approach to screen for drug candidates that could be used as an alternative to taxanes, based on a 86 genes signature which could distinguish between low-grade and high-grade tumors. We extracted from the NCI60 panel databases the expression profiles of the 86 genes across 60 human tumor cell lines and the corresponding in vitro cytotoxicity data of 152 drugs and looked for correlation between their expression level and cell sensitivity to each of these drugs. Results: Among the 86 genes, we identified 9 genes (PCCB, SHMT2, DPM1, RHOT2, RPL13, CD59, EIF4AI, CDKN2C, JUN) for which expression levels across the 60 cell lines was significantly correlated (p< 0.05) to oxaliplatin but not to cisplatin sensitivity. This signature was validated at the functional level since repression of each of these genes conferred a significant change in the sensitivity of PCa cell lines to oxaliplatin but not cisplatin. Conclusions: Our results provide a proof of concept that gene expression signature specific from high grade PCa could be used for the identification of alternative therapies to taxanes. They could also be used to select patients for further clinical trials and as predictive markers of response to these drugs, which represents a further step forward towards personalized therapy of PCa.

Identification of synthetic lethal interactions with the BRAF oncogene in colorectal cancer.

2013 ◽  
Vol 31 (4_suppl) ◽  
pp. 403-403
Author(s):  
Loredana Vecchione ◽  
Valentina Gambino ◽  
Giovanni d'Ario ◽  
Sun Tian ◽  
Iris Simon ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Lines ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Gene Expression Signature ◽  
Gene Signature ◽  
Braf V600e ◽  
P Value ◽  
Synthetic Lethal ◽  
Braf Mutant

403 Background: Approximately 8-15% of colorectal (CRC) patients carry an activating mutation in BRAF. This CRC subtype is associated with poor outcome and with resistance, both to chemotherapeutic treatments and to tailored drugs. We recently showed that BRAF (V600E) colon cancers (CCs) have a characteristic gene expression signature (1, 2) which is found also in subsets of KRAS mutant and KRAS-BRAF wild type (WT2) tumors. Tumors having this gene signature, referred as “BRAF-like”, have a similar poor prognosis irrespective of the presence of the BRAF (V600E) mutation. By using a shRNA-based genetic screen in BRAF mutant CC cell lines we aimed to identify genes and pathways necessary for survival and growth of BRAFmutant CC. Such studies may reveal additional targets for therapy and potentially provide new biomarkers for patient stratification Methods: We identified 363 genes that are selectively overexpressed in BRAF mutant tumors as compared to WT2 type tumors, based on gene expression profiles of the PETACC3 (1) and Agendia (2) datasets. The TRC human genome-wide shRNA collection (TRC-Hs1.0) was used to generate a 1815 hairpins sub-library targeting those identified genes (BRAF library). BRAF(V600E) CC cell lines were infected with the BRAF library and screened for shRNAs that cause lethality. LIM1215 CC cell line (WT2) was used as a control. Cells stably expressing the shRNA library were cultured for 13 days, after which shRNAs were recovered by PCR. Deep sequencing was applied to determine the specific depletion of shRNA in BRAF(V600E) cells as compared to LIM1215 cells Results: Candidate genes were identified by using following filtering criteria: depletion in BRAF(V600E) cells by at least 50% and depletion in BRAF(V600E) cells 1, 5-fold higher than in control cells with the corresponding p-value to be ≤ 0.1. A total of 34 genes met our criteria of which 6 genes were presented with more than one hairpin and were concordant across the cell lines selected for validation. Conclusions: We identified candidate synthetic lethal genes in BRAF mutant CC cell lines. Functional analysis is ongoing. Data will be presented. References 1. J Clin Oncol 2012 Apr 20;30(12):1288-9 2. Gut (2012). doi:10.1136/gutjnl-2012-302423

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